Project Summary Technological advances have enabled biologists to categorize entire omes? genomes, but also transcriptomes, proteomes, metabolomes, etc., down to the level of individual cells. Systems Biology is an ascendant branch of biology, with the goal of understanding the interactions between the molecular components of an organism, and how those interactions function to build, operate, and maintain the organism in the context of its environment. The number of such interactions is vast, but experience suggests that there will be underlying consistent rules. A potential way forward is to scrutinize features of a large system ? a transcriptome, for example ? for consistent signatures of natural selection. A powerful way to reveal the signature of natural selection is to compare the genetic variation introduced by mutation to the standing genetic variation in the population. That is because the standing genetic variation has been scrutinized by natural selection. A consistent discrepancy between the genetic variation introduced by spontaneous mutation and the standing genetic variation present in a species is an unmistakable signature of natural selection. In turn, identifying some feature of an organism that is demonstrably under natural selection is prima facie evidence that the feature has a significant biological function, even if the function is not immediately obvious. The raw material for systems biologists is a (large) set of measures of the abundance of individual transcripts, proteins, metabolites, etc. It seems likely that in most cases the mutational target of an individual transcript, etc., will be small. If the mutational target is small, many genomes must be screened to provide a reliable characterization of the mutational process responsible for producing genetic variation in the trait of interest. The goal of the proposed work is to construct a large set of replicate populations of the model nematode Caenorhabditis elegans that have evolved under minimal natural selection, thereby allowing all but the most highly deleterious mutations to accumulate as if they are invisible to natural selection. Ultimately, the set of mutation accumulation lines are expected to harbor approximately 100,000 spontaneous mutations. C. elegans provides major advantages over other animal models in this context, the most important being that nematodes can be easily and reliably cryopreserved. The resource will therefore be durable, and experiments can be done on the (nearly) exact same genetic stock for decades hence. The genomes of the lines will be sequenced, thereby allowing researchers to associate genotypes with their specific traits of interest. Both the lines themselves and the genome sequences will be made immediately available as a community resource. Systems Biology is inherently concerned with interactions between genes, and between genes and the environment. Model organisms such as C. elegans are especially valuable in that regard because genotypes and environments can be both carefully controlled, neither of which is possible with human subjects.